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Search indexed NASA NTRS and DOE OSTI research on propulsion, heat transfer, battery materials and energy systems. Follow report and document links to the original sources.

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At least 19 records
All sourcesNASA NTRSDOE OSTI
DOE OSTI2026

Compartmental Control of UDP-Araf Supply: Golgi Lumen UDP-Arabinopyranose Mutase Depletes Plant Cell Wall Araf Linkages

L-Arabinofuranose (Araf) is a major constituent of plant cell wall polysaccharides. UDP-Araf is generated from UDP-L-arabinopyranose (UDP-Arap) by a UDP-Arap mutase (UAM) on the cytosolic face of the Golgi and then imported into the Golgi lumen for utilization by arabinosyltransferases (AraTs). Yet, a substantial fraction of UDP-Arap is synthesized in the Golgi lumen by the UDP-xylose 4-epimerase (UXE), creating a topological paradox where luminal UDP-Arap must be exported to the cytosol for conversion to UDP-Araf by UAM and then re-imported into the Golgi lumen. To test the functional significance of this compartmentation, we mislocalized Arabidopsis (Arabidopsis thaliana) UAM1 to the Golgi lumen by fusing it to the UXE1 transmembrane domain. The fusion protein was present in the Golgi, and protease-protection assays on microsomes supported its luminal orientation. In wild-type and uam1 plants, Golgi-targeted UAM1 reduced total cell wall Arabinose without consistent changes in other monosaccharides. Linkage analyses showed that this decrease reflects selective loss of Araf residues, whereas Arap remain largely unchanged, consistent with reduced UDP-Araf availability for luminal AraTs. Under salt stress, expression of Golgi-targeted UAM1 in uam1 caused root-growth inhibition and maturation-zone swelling that exogenous L-Ara failed to rescue. These results support a hypothesis that the strong thermodynamic bias of the UAM reaction toward UDP-Arap requires spatial separation of UDP-Arap production from UDP-Araf formation to sustain UDP-Araf flux into Golgi arabinosylation and avoid futile intraluminal back-conversion.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

UDP-glucuronic acid decarboxylase in alfalfa: a target to improve ruminal digestibility of stems

Alfalfa (Medicago sativa) has a high nutritional value, but poor digestibility of the stems limits its value as an energy source in ruminant diets. Xylan and lignin negatively affect cell wall digestibility, whereas pectins have high digestibility in the rumen. In plants, UDP-xylose synthase (UXS) catalyses the decarboxylation of UDP-glucuronic acid to form UDP-xylose in an irreversible step that is key for xylan synthesis. Here, we functionally characterized two UXS genes in alfalfa, namely MsaUXS2 and MsaUXS4, and investigated their impact on ruminal digestibility. Both genes are more highly expressed in stems than leaves, and the enzymes have UDP-glucuronic acid decarboxylase activity in vitro. Silencing of MsaUXS2 and MsaUXS4 via RNAi altered plant growth and resulted in a 40% decrease in xylose, a 115% increase in arabinose, and a 60% increase in galacturonic acid in the polysaccharide matrix as well as a 20% decrease in lignin in the cell wall. Together, our results show a major role for UXS2 and UXS4 in xylan synthesis and secondary cell wall deposition in alfalfa. Additionally, in vitro rumen digestibility assays for the silenced lines had on average 30% increased gas production at 24 h, demonstrating the potential of targeting UXS genes to increase stem digestibility.

UDP-xylose synthase↗
DOE OSTI2020

Structural basis of the UDP-diacylglucosamine pyrophosphohydrolase LpxH inhibition by sulfonyl piperazine antibiotics

The UDP-2,3-diacylglucosamine pyrophosphate hydrolase LpxH is an essential lipid A biosynthetic enzyme that is conserved in the majority of gram-negative bacteria. It has emerged as an attractive novel antibiotic target due to the recent discovery of an LpxH-targeting sulfonyl piperazine compound (referred to as AZ1) by AstraZeneca. However, the molecular details of AZ1 inhibition have remained unresolved, stymieing further development of this class of antibiotics. In this paper we report the crystal structure of Klebsiella pneumoniae LpxH in complex with AZ1. We show that AZ1 fits snugly into the L -shaped acyl chain-binding chamber of LpxH with its indoline ring situating adjacent to the active site, its sulfonyl group adopting a sharp kink, and its N -CF 3 –phenyl substituted piperazine group reaching out to the far side of the LpxH acyl chain-binding chamber. Intriguingly, despite the observation of a single AZ1 conformation in the crystal structure, our solution NMR investigation has revealed the presence of a second ligand conformation invisible in the crystalline state. Together, these distinct ligand conformations delineate a cryptic inhibitor envelope that expands the observed footprint of AZ1 in the LpxH-bound crystal structure and enables the design of AZ1 analogs with enhanced potency in enzymatic assays. These designed compounds display striking improvement in antibiotic activity over AZ1 against wild-type K. pneumoniae , and coadministration with outer membrane permeability enhancers profoundly sensitizes Escherichia coli to designed LpxH inhibitors. Remarkably, none of the sulfonyl piperazine compounds occupies the active site of LpxH, foretelling a straightforward path for rapid optimization of this class of antibiotics.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

Temporal Galactose‐Manganese Feeding in Fed‐Batch and Perfusion Bioreactors Modulates UDP‐Galactose Pools for Enhanced mAb Glycosylation Homogeneity

ABSTRACT Monoclonal antibodies (mAbs) represent a majority of biotherapeutics in the market today. These glycoproteins undergo posttranslational modifications, such as N‐linked glycosylation, that influence the structural & functional characteristics of the antibody. Glycosylation is a heterogenous posttranslational modification that may influence therapeutic glycoprotein stability and clinical efficacy, which is why it is often considered a critical quality attribute (CQA) of the mAb product. While much is known about the glycosylation pathways of Chinese Hamster Ovary (CHO) cells and how cell culture chemical modifiers may influence the N‐glycosylation profile of the final product, this knowledge is often based on the final cumulative glycan profile at the end of the batch process. Building a temporal understanding of N‐glycosylation and how mAb glycoform composition responds to real‐time changes in the biomanufacturing process will help build integrated process models that may allow for glycosylation control to produce a more homogenous product. Here, we look at the effect of specific nutrient feed media additives (e.g., galactose, manganese) and feeding times on the N‐glycosylation pathway to modulate N‐glycosylation of a Herceptin biosimilar mAb (i.e., Trastuzumab). We deploy the N‐GLYcanyzer process analytical technology (PAT) to monitor glycoforms in near real‐time for bench‐scale bioprocesses operated in both fed‐batch and perfusion modes to build an understanding of how temporal changes in mAb N‐glycosylation are dependent on specific media additives. We find that Trastuzumab terminal galactosylation is sensitive to media feeding times and intracellular nucleotide sugar pools. Temporal analysis reveals an increased desirable production of single and double galactose‐occupied glycoforms over time under glucose‐starved fed‐batch cultures. Comparable galactosylation profiles were also observed between fed‐batch (nutrient‐limited) and perfusion (non‐nutrient‐limited) bioprocess conditions. In summary, our results demonstrate the utility of real‐time monitoring of mAb glycoforms and feeding critical cell culture nutrients under fed‐batch and perfusion bioprocessing conditions to produce higher‐quality biologics.

Biotechnology & Applied Microbiology↗
DOE OSTI2022

Bantamweight Badger UDP support (BWUDP) v1.0

Software networking library to allow an FPGA ethernet core (such as Badger: https://gitlab.lbl.gov/hdl-libraries/bedrock/-/tree/master/badger) to be used in a soft-core processor or microcontroller.

Russo, Lucas↗
DOE OSTI2023

Validation of a metabolite–GWAS network for Populus trichocarpa family 1 UDP-glycosyltransferases

Metabolite genome-wide association studies (mGWASs) are increasingly used to discover the genetic basis of target phenotypes in plants such as Populus trichocarpa , a biofuel feedstock and model woody plant species. Despite their growing importance in plant genetics and metabolomics, few mGWASs are experimentally validated. Here, we present a functional genomics workflow for validating mGWAS-predicted enzyme–substrate relationships. We focus on uridine diphosphate–glycosyltransferases (UGTs), a large family of enzymes that catalyze sugar transfer to a variety of plant secondary metabolites involved in defense, signaling, and lignification. Glycosylation influences physiological roles, localization within cells and tissues, and metabolic fates of these metabolites. UGTs have substantially expanded in P. trichocarpa , presenting a challenge for large-scale characterization. Using a high-throughput assay, we produced substrate acceptance profiles for 40 previously uncharacterized candidate enzymes. Assays confirmed 10 of 13 leaf mGWAS associations, and a focused metabolite screen demonstrated varying levels of substrate specificity among UGTs. A substrate binding model case study of UGT-23 rationalized observed enzyme activities and mGWAS associations, including glycosylation of trichocarpinene to produce trichocarpin, a major higher-order salicylate in P. trichocarpa. We identified UGTs putatively involved in lignan, flavonoid, salicylate, and phytohormone metabolism, with potential implications for cell wall biosynthesis, nitrogen uptake, and biotic and abiotic stress response that determine sustainable biomass crop production. Our results provide new support for in silico analyses and evidence-based guidance for in vivo functional characterization.

59 BASIC BIOLOGICAL SCIENCES↗
NASA NTRS2017

Method and Apparatus for Processing UDP Data Packets

A method and apparatus for processing a plurality of data packets. A data packet is received. A determination is made as to whether a portion of the data packet follows a selected digital recorder standard protocol based on a header of the data packet. Raw data in the data packet is converted into human-readable information in response to a determination that the portion of the data packet follows the selected digital recorder standard protocol.

Murphy, Brandon M.↗
DOE OSTI2024

Characterization of the Neurospora crassa Galactosaminogalactan Biosynthetic Pathway

The Neurospora crassa genome has a gene cluster for the synthesis of galactosaminogalactan (GAG). The gene cluster includes the following: (1) UDP-glucose-4-epimerase to convert UDP-glucose and UDP-N-acetylglucosamine to UDP-galactose and UDP-N-acetylgalactosamine (NCU05133), (2) GAG synthase for the synthesis of an acetylated GAG (NCU05132), (3) GAG deacetylase (/NCW-1/NCU05137), (4) GH135-1, a GAG hydrolase with specificity for N-acetylgalactosamine-containing GAG (NCU05135), and (5) GH114-1, a galactosaminidase with specificity for galactosamine-containing GAG (NCU05136). The deacetylase was previously shown to be a major cell wall glycoprotein and given the name of NCW-1 (non-GPI anchored cell wall protein-1). Characterization of the polysaccharides found in the growth medium from the wild type and the GAG synthase mutant demonstrates that there is a major reduction in the levels of polysaccharides containing galactosamine and N-acetylgalactosamine in the mutant growth medium, providing evidence that the synthase is responsible for the production of a GAG. The analysis also indicates that there are other galactose-containing polysaccharides produced by the fungus. Phenotypic characterization of wild-type and mutant isolates showed that deacetylated GAG from the wild type can function as an adhesin to a glass surface and provides the fungal mat with tensile strength, demonstrating that the deacetylated GAG functions as an intercellular adhesive. The acetylated GAG produced by the deacetylase mutant was found to function as an adhesive for chitin, alumina, celite (diatomaceous earth), activated charcoal, and wheat leaf particulates.

59 BASIC BIOLOGICAL SCIENCES↗
NASA NTRS2018

Use of Nack Oriented Reliable Multicast (NORM) Protocol for Transport of Spacecraft Telemetry in Ground Networks

Frequently it is necessary to distribute spacecraft telemetry to multiple destinations on Internet Protocol (IP) networks. There are various methods of delivering this data, but when the data is high rate and there are many destinations IP multicast provides the best solution from a network perspective. However, IP multicast only supports the User Datagram Protocol (UDP) which is connectionless and unreliable. This nature of UDP and by extension the low priority treatment of UDP packets by some network hardware creates a difficult environment for distribution of high rate telemetry to a number of recipients over IP networks. NACK (Negative ACKnowledgment) Oriented Reliable Multicast (NORM) is defined by IETF (Internet Engineering Task Force) RFC (Request for Comments) 5740 in 2009 by B. Adamson et al. NORM provides a mechanism of retransmission and or forward error correction for UDP packets, potentially overcoming the limits of UDP for the transmission of spacecraft telemetry. Marshall Space Flight Center’s (MSFC) Huntsville Operations Support Center (HOSC) manages the delivery of the International Space Station (ISS) payload science and health data. The ISS IP Ground Routed (IIGoR) network provides the delivery of the data from the ground stations to the control centers. HOSC personnel have been experimenting with the use of NORM on the link between the ground stations and the front end processor equipment in Huntsville, AL.

Baldwin, Patrick A.↗
DOE OSTI2021

Comparison of human poly-N-acetyl-lactosamine synthase structure with GT-A fold glycosyltransferases supports a modular assembly of catalytic subsites

Poly-N-acetyl-lactosamine (poly-LacNAc) structures are composed of repeating [-Galβ(1,4)-GlcNAcβ(1,3)-]n glycan extensions. They are found on both N- and O-glycoproteins and glycolipids and play an important role in development, immune function, and human disease. The majority of mammalian poly-LacNAc is synthesized by the alternating iterative action of β1,3-N-acetylglucosaminyltransferase 2 (B3GNT2) and β1,4-galactosyltransferases. B3GNT2 is in the largest mammalian glycosyltransferase family, GT31, but little is known about the structure, substrate recognition, or catalysis by family members. Here we report the structures of human B3GNT2 in complex with UDP:Mg 2+ and in complex with both UDP:Mg 2+ and a glycan acceptor, lacto-N-neotetraose. The B3GNT2 structure conserves the GT-A fold and the DxD motif that coordinates a Mg 2+ ion for binding the UDP-GlcNAc sugar donor. The acceptor complex shows interactions with only the terminal Galβ(1,4)-GlcNAcβ(1,3)- disaccharide unit, which likely explains the specificity for both N- and O-glycan acceptors. Modeling of the UDP-GlcNAc donor supports a direct displacement inverting catalytic mechanism. Comparative structural analysis indicates that nucleotide sugar donors for GT-A fold glycosyltransferases bind in similar positions and conformations without conserving interacting residues, even for enzymes that use the same donor substrate. In contrast, the B3GNT2 acceptor binding site is consistent with prior models suggesting that the evolution of acceptor specificity involves loops inserted into the stable GT-A fold. These observations support the hypothesis that GT-A fold glycosyltransferases employ coevolving donor, acceptor, and catalytic subsite modules as templates to achieve the complex diversity of glycan linkages in biological systems.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗
DOE OSTI2025

Identification and characterization of mono- and bifunctional galactan synthases in the pediatric pathogen Kingella kingae

The emerging pediatric pathogen Kingella kingae elaborates a lipopolysaccharide (LPS) that is extended with a galactofuranose homopolymer called galactan, which is a key virulence determinant that contributes to resistance to complement-mediated and neutrophil-mediated killing. Previous work has demonstrated that the pamABCDE locus is required for galactan synthesis. In this study, mutational studies suggested that the pamC gene product is a UDP-galactofuranose (Galf) transferase and is the galactan synthase. Analysis of genome sequence data revealed two distinct pamC alleles designated pamC1 and pamC2, which correlate with the two galactan structures in K. kingae. Examination of isogenic mutants expressing either pamC1 or pamC2 demonstrated that the pamC alleles are the determinants of galactan structure. Experiments with recombinant PamC1 and PamC2 in vitro established that these proteins are galactan synthases capable of extending synthetic Galf disaccharide acceptors in the presence of UDP-Galf. Homology analysis identified critical amino acids that are essential for PamC1 and PamC2 enzymatic activity both in vitro and in K. kingae. Structural analysis of the in vitro-modified synthetic acceptors implicated PamC1 as a monofunctional enzyme capable of generating a β-(1 → 5) Galf linkage and PamC2 as a bifunctional enzyme capable of generating β-(1 → 3) and β-(1 → 6) Galf linkages. This study advances our understanding of the GT2 family of UDP-galactofuranosyltransferases.

60 APPLIED LIFE SCIENCES↗
DOE OSTI2023

Deciphering triterpenoid saponin biosynthesis by leveraging transcriptome response to methyl jasmonate elicitation in Saponaria vaccaria

Abstract Methyl jasmonate (MeJA) is a known elicitor of plant specialized metabolism, including triterpenoid saponins. Saponaria vaccaria is an annual herb used in traditional Chinese medicine, containing large quantities of oleanane-type triterpenoid saponins with anticancer properties and structural similarities to the vaccine adjuvant QS-21. Leveraging the MeJA-elicited saponin biosynthesis, we identify multiple enzymes catalyzing the oxidation and glycosylation of triterpenoids in S. vaccaria . This exploration is aided by Pacbio full-length transcriptome sequencing and gene expression analysis. A cellulose synthase-like enzyme can not only glucuronidate triterpenoid aglycones but also alter the product profile of a cytochrome P450 monooxygenase via preference for the aldehyde intermediate. Furthermore, the discovery of a UDP-glucose 4,6-dehydratase and a UDP-4-keto-6-deoxy-glucose reductase reveals the biosynthetic pathway for the rare nucleotide sugar UDP- d -fucose, a likely sugar donor for fucosylation of plant natural products. Our work enables the production and optimization of high-value saponins in microorganisms and plants through synthetic biology approaches.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

Multiple Arabidopsis galacturonosyltransferases synthesize polymeric homogalacturonan by oligosaccharide acceptor‐dependent or de novo synthesis

SUMMARY Homogalacturonan (HG), the most abundant pectic glycan, functions as a cell wall structural and signaling molecule essential for plant growth, development and response to pathogens. HG exists as a component of pectic homoglycans, heteroglycans and glycoconjugates. HG is synthesized by members of the GALACTURONOSYLTRANSFERASE (GAUT) family. UDP‐GalA‐dependent homogalacturonan:galacturonosyltransferase (HG:GalAT) activity has previously been demonstrated for GAUTs 1, 4 and 11, as well as the GAUT1:GAUT7 complex. Here, we show that GAUTs 10, 13 and 14 are also HG:GalATs and that GAUTs 1, 10, 11, 13, 14 and 1:7 synthesize polymeric HG in vitro . Comparison of the in vitro HG:GalAT specific activities of the heterologously‐expressed proteins demonstrates GAUTs 10 and 11 with the lowest, GAUT1 and GAUT13 with moderate, and GAUT14 and the GAUT1:GAUT7 complex with the highest HG:GalAT activity. GAUT13 and GAUT14 are also shown to de novo synthesize (initiate) HG synthesis in the absence of exogenous HG acceptors, an activity previously demonstrated for GAUT1:GAUT7. The rate of de novo HG synthesis by GAUT13 and GAUT14 is similar to their acceptor dependent HG synthesis, in contrast to GAUT1:GAUT7 for which de novo synthesis occurred at much lower rates than acceptor‐dependent synthesis. The results suggest a unique role for de novo HG synthesis by GAUTs 13 and 14. The reducing end of GAUT13‐ de novo‐ synthesized HG has covalently attached UDP, indicating that UDP‐GalA serves as both a donor and acceptor substrate during de novo HG synthesis. The functional significance of unique GAUT HG:GalAT catalytic properties in the synthesis of different pectin glycan or glycoconjugate structures is discussed.

Engle, Kristen A.↗
NASA NTRS2006

Improving UDP/IP Transmission Without Increasing Congestion

Datagram Retransmission (DGR) is a computer program that, within certain limits, ensures the reception of each datagram transmitted under the User Datagram Protocol/Internet Protocol. [User Datagram Protocol (UDP) is considered unreliable because it does not involve a reliability-ensuring connection-initiation dialogue between sender and receiver. UDP is well suited to issuing of many small messages to many different receivers.] Unlike prior software for ensuring reception of UDP datagrams, DGR does not contribute to network congestion by retransmitting data more frequently as an ever-increasing number of messages and acknowledgements is lost. Instead, DGR does just the opposite: DGR includes an adaptive timeout-interval- computing component that provides maximum opportunity for reception of acknowledgements, minimizing retransmission. By monitoring changes in the rate at which message-transmission transactions are completed, DGR detects changes in the level of congestion and responds by imposing varying degrees of delay on the transmission of new messages. In addition, DGR maximizes throughput by not waiting for acknowledgement of a message before sending the next message. All DGR communication is asynchronous, to maximize efficient utilization of network connections. DGR manages multiple concurrent datagram transmission and acknowledgement conversations.

Burleigh, Scott↗
NASA NTRS2016

Next Generation Flight Displays Using HTML5

The Human Integrated Vehicles and Environments (HIVE) lab at Johnson Space Center (JSC) is focused on bringing together inter-disciplinary talent to design and integrate innovative human interface technologies for next generation manned spacecraft. As part of this objective, my summer internship project centered on an ongoing investigation in to building flight displays using the HTML5 standard. Specifically, the goals of my project were to build and demo "flight-like" crew and wearable displays as well as create a webserver for live systems being developed by the Advanced Exploration Systems (AES) program. In parallel to my project, a LabVIEW application, called a display server, was created by the HIVE that uses an XTCE (XML (Extensible Markup Language) Telemetry and Command Exchange) parser and CCSDS (Consultative Committee for Space Data System) space packet decoder to translate telemetry items sent by the CFS (Core Flight Software) over User Datagram Protocol (UDP). It was the webserver's job to receive these UDP messages and send them to the displays. To accomplish this functionality, I utilized Node.js and the accompanying Express framework. On the display side, I was responsible for creating the power system (AMPS) displays. I did this by using HTML5, CSS and JavaScript to create web pages that could update and change dynamically based on the data they received from the webserver. At this point, I have not started on the commanding, being able to send back to the CFS, portion of the displays but hope to have this functionality working by the completion of my internship. I also created a way to test the webserver's functionality without the display server by making a JavaScript application that read in a comma-separate values (CSV) file and converted it to XML which was then sent over UDP. One of the major requirements of my project was to build everything using as little preexisting code as possible, which I accomplished by only using a handful of JavaScript libraries. As a side project, I created a model of the HIVE lab and Building 29 using SketchUp. I obtained the floorplans of the building from the JSC Geographic Information Systems (GIS), which were computer-aided design (CAD) files, and imported them into SketchUp. I then took those floorplans and created a 3D model of the building from them. Working in conjunction with the Hybrid Reality lab in Building 32, the SketchUp model was imported into Unreal Engine for use with the HTC Vive. Using the Vive, I was able to interact with the model I created in virtual reality (VR). The purpose of this side project was to be able to visualize potential lab layouts and mockup designs as they are in development in order to finalize design decisions. Pending approval, the model that I created will be used in the Build-As-You-Test: Can Hybrid Reality Improve the SE/HSI Design Process project in the fall. Getting the opportunity to work at NASA has been one of the most memorable experiences of my life. Over the course of my internship, I improved my programming and web development abilities substantially. I will take all the skills and experiences I have had while at NASA back to school with me in the fall and hope to pursue a career in the aerospace industry after graduating in the spring.

Greenwood, Brian↗
DOE OSTI2026

A characterization of recombinant Arabidopsis FRIABLE1 (FRB1) reveals robust rhamnogalacturonan-I rhamnosyltransferase activity and critical catalytic residues

Plant cell walls are glycan-rich extracellular matrices that fundamentally impact essential cellular processes, such as growth, adhesion, and cell shape acquisition. Understanding plant cell wall glycans requires the identification and characterization of the biosynthetic enzymes that produce these polymers. Most successful in vitro protein expression studies of plant cell wall glycosyltransferases have relied on insect, fungal/yeast, or human cell expression systems, whereas prokaryotic expression systems have been generally unsuccessful. Here, we show that Arabidopsis FRIABLE1 (FRB1)/rhamnogalacturonan-I rhamnosyltransferase 8 (RRT8) can be produced in Escherichia coli RosettaGami2 cells as N-terminal maltose-binding protein fusion proteins containing C-terminal 6X-His-tags. We also report the catalytic constants of FRB1/RRT8 with apparent K M and K cat values of 226 μM and 33 min -1 for UDP-Rhamnose and 117 μM and 28.7 min -1 for rhamnogalacturonan-I (RG-I), respectively. We examine the catalytic activities of mutated FRB1/RRT8 proteins based on an AlphaFold 3-generated FRB1/RRT8 protein structural model with a virtually docked UDP-Rha donor. Enzymatic characterization of the mutated and wildtype FRB1/RRT8 protein confirmed that mutation of predicted catalytic site amino acid residues resulted in a 20-fold reduction in RRT activity. FRB1 also robustly polymerizes RG-I in combination with RG-I galacturonosyltransferase 1. These results show how a robust E. coli expression system combined with artificial intelligence tools can be used to increase understanding of plant cell wall glycosyltransferase structure and function.

glycosyltransferase↗
DOE OSTI2023

Insights into substrate coordination and glycosyl transfer of poplar cellulose synthase-8

Cellulose is an abundant cell wall component of land plants. It is synthesized from UDP-activated glucose molecules by cellulose synthase, a membrane-integrated processive glycosyltransferase. Cellulose synthase couples the elongation of the cellulose polymer with its translocation across the plasma membrane. Here, we present substrate- and product-bound cryogenic electron microscopy structures of the homotrimeric cellulose synthase isoform-8 (CesA8) from hybrid aspen (poplar). UDP-glucose binds to a conserved catalytic pocket adjacent to the entrance to a transmembrane channel. The substrate’s glucosyl unit is coordinated by conserved residues of the glycosyltransferase domain and amphipathic interface helices. Site-directed mutagenesis of a conserved gating loop capping the active site reveals its critical function for catalytic activity. Molecular dynamics simulations reveal prolonged interactions of the gating loop with the substrate molecule, particularly across its central conserved region. These transient interactions likely facilitate the proper positioning of the substrate molecule for glycosyl transfer and cellulose translocation.

59 BASIC BIOLOGICAL SCIENCES↗